The long-term immune effects of cigarette smoking especially in the lung remain a poorly understood area of research. We have demonstrated that antigen presenting cells (APCs) are highly activated in the lungs of smokers with emphysema, and mice exposed to smoke increase pathogenic APCs in the lungs that is critical for T helper (Th) 1, and Th17 inflammation and emphysema development. How tobacco smoke activates APCs and the downstream molecular mechanisms responsible for lung tissue injury remain unknown. In search of endogenous molecules that could promote maladaptive lung immune responses, we found elevated plasma concentrations of complement protein 3 (C3) in smokers with emphysema, and show that neutrophil elastase (NE) and MMP12 cleave and release its active fragments. We have also found that C3-/- mice exposed to cigarette smoke have reduced lung inflammation and attenuated emphysema. Interestingly this phenotype in C3-/- mice was marked by immature lung APCs and preserved expression of another complement protein C1q, lack of which has been associated with autoimmune inflammatory diseases. Further, treatment of mice with C1q reduced lung inflammation and emphysema while inhibition of C1q with siRNA increased their ability to induce Th17 effector cells. Collectively, these new critical observations point to an immune modulatory role for complement proteins in emphysema; the clinical significance of our proposed studies includes discovering novel and effective non-immune compromising complement-based therapeutics for treatment of smoke- induced lung diseases. Our three Aims are: Aim 1. Identify mechanism(s) activating complement and stimulating pathogenic APCs in lungs exposed to inhale cigarette smoke. Hypothesis: C3a and/or C5a are required for efficient activation of pathogenic APCs in the lungs in response to cigarette smoke. This work will be accomplished by i) investigating the mechanism(s) for C3 mediated APC activation and lung immune phenotype of wild type, C3aR-/-, and C5aR-/- mice exposed to chronic smoke; using the same model, the efficacy of C3 inhibitor (e.g. CR2-fH) will be examined in vivo ii) In human translational studies using siRNA the function of C3a and C5a signaling in APC-mediated activation of CD4 T cells will be explored. Aim 2. Determine the effect of C1q on lung APCs in cigarette smoke-induced lung inflammation. Hypothesis: Reduction of C1q in APCs of mice exposed to smoke stimulates Th1/Th17 mediated inflammation and worsens emphysema whereas production of C1q by APC reduces inflammatory responses that result in emphysema. We will explore the role of C1q using C1q-/- mice and specific C1q deficiency in APCs (CD11ccre/C1qflx/flx mice) in a model of smoke induced emphysema. Complementary studies using loss of function (siRNA) and gain of function (add back C1q protein) will be used to further determine the role of C1q modulation in APCs in human emphysema. Aim 3. Determine the mechanism(s) for the crosstalk between C3 and C1q in lung APCs of smoke exposed mice. Hypotheses: C1q promotes development of tolerogenic lung APCs. Using transgenic mice expressing GFP under Foxp3 promoter (Foxp3GFP) the role of C3, C3aR and C5aR in C1q mediated induction of T cell tolerance, and production of inhibitory cytokines will be examined in response to cigarette smoke. This proposal is based on our human translational studies that are focused on the role of complement proteins in promoting development of autoreactive immune responses in the lungs. Successful completion of the objectives should provide a strong rationale for the use of novel and effective non-immune compromising complement-based therapeutics for treatment of smoke-induced lung diseases.